My lab is interesting in sorting one and two cell embryos. We have tried following the nature methods paper published a few years ago, using the FACsVantage. The problem is, once we got it to work on the FACSVantage, our core facility sold the equipment and bought a FACSAria, which we have been unable to reproduce the result with. I am wondering if anyone is using the FACSAria and getting the 98% pure population of 1-2 cell embryos that is possible to get on the Vantage. Or if anyone knows a core facility that still has a functional Vantage that I can use. Thanks!
Hey,
we are using AriaIII and have a lot better sorting performance than with the Vantage. The machine is faster, runs more stably with the 100µm nozzle and also yields a very pure population of embryos already after the first sort.
best,
Marlon
Thanks for the response. Have you ever used the AriaII? That is what we have access to and we find it much slower and virtually impossible to find the population of GFP positive cells that we saw on the Vantage. Thanks
Yes, I also tested the ariaII, it also worked quite well. Finding the GFP+ cells in my impression it pretty much depends on the ‘autofluorescence’ channel (e.g. red fluorescence) which you choose and the right compensation.
best,
marlon
Marlon,
Are you still using a one hour methanol fixation? If possible could you send us the compensation parameters? Also would it be possible for you to send me your TX189 strain. You can email me at: tmedwar@emory.edu for the address and fedex number if you are willing to do that. Thanks!
I have the strain from R.Lin Lab but it’s GFP fluorescence is comparable to the strain that R.Lin deposited at CGC. Compensation settings are pretty much dependent on the lasers you use and are quite variable. Send me a picture of the profile you see and I can look over it if you want:
marlon.stoeckius@mdc-berlin.de
Best,
Marlon